Mannich Reaction With Pyridoxal 5′‐Phosphate Dependent Decarboxylative Aldolase for Synthesis of Noncanonical α ‐Amino Acids

Functionalized benzosultam and sulfonamide derivatives represent privileged pharmacophores in medicinal chemistry. However, noncanonical amino acids (ncAAs) featuring benzosultam side chains remain underexplored for peptide therapeutics, despite significant advances in peptide drug development. Herein, we report the promiscuous activity of the PLP‐dependent decarboxylative aldolase UstD in catalyzing β ‐Mannich reactions. An engineered variant, UstD2.0 S60A , was developed and applied to synthesize 23 enantioenriched ncAAs, achieving yields up to 96%, a total turnover number (TTN) of 7500, and excellent stereoselectivity (>95:5 dr, >99:1 er). The reaction is scalable to gram quantities for preparative applications. This study facilitates the future use of benzosultam‐containing ncAAs in peptide chemistry and highlights the opportunity of engineering aldolases for non‐natural Mannich reaction to access chiral amine products.